This invention relates in general to solid-phase immunoassays and, more particularly, to a method for drying mammalian cells adsorbed to a solid-phase support for use in solid-phase immunoassays and articles incorporating same.
Solid-phase immunological assay procedures have been developed to provide sensitive and specific immunological assays that are more convenient and easier for lab technicians to perform. In a solid-phase immunoassay, one component of the immunological reaction, either antigen or antibody, is immobilized onto the surface of a solid-phase support. The antigen or antibody may be immobilized directly onto the solid-phase support by a variety of methods known to those skilled in the art. In addition to antigens or antibodies, whole cells such as erythrocytes, leukocytes, lymphocytes, platelets or other mammalian cells can be immobilized to the surface of a solid-phase support for immunological assays specific for such cells. Each of these types of cells present antigens on its cell surface which can be used to detect antibodies in a biological fluid sample to be assayed. Also, specific antigens or antibodies can be adsorbed to the surface of the cells for other types of immunological assays. Thus, immunoassays involving cells of the type described above have been developed to determine the presence of antigens or antibodies in assays for blood groups and infectious disease agents, especially those directly affecting the cellular components of blood.
When whole cells are utilized in solid-phase immunoassays, present techniques used to fix these cells to the solid-phase support employ chemical agents which crosslink functional groups on the surface of the cells to functional groups on the surface of the solid-phase support. Typical chemical agents used for this purpose are glutaraldehyde and dimethyl suberimidate. The primary problem associated with the use of these agents for immunoassays involving whole mammalian cells is that while they fix the cells to the support, they also block some reactive groups which are responsible for antigenic activity. Due to the enormous complexity and multitude of antigenic determinants found upon the surface of cells, no single chemical crosslinking agent has been found which does not affect the antigenicity of the cell membrane. In those assays where only a single or only a few antigenic determinants must be maintained, chemical cross-linking agents can be utilized effectively. If all of the antigenic determinants must be maintained, however, as in the case of assays utilizing red cell monolayers used to detect irregular antibodies in patient sera, chemical cross-linking agents cannot be used since they block some of the potential reactive groups.
Whole mammalian cells may also be fixed by agents which draw water out of the cell. Typical agents that draw water out of or replace water within cells are methanol, acetone and dimethyl sulfoxide. Methanol and acetone have been used to fix tissue slices for microscopic examination, but are not applicable for use with immobilized monolayers of cells such as erythrocytes, lymphocytes, or platelets because they tend to destroy antigens on the surface of the cells, especially red blood cells. Additional problems with chemicals like methanol and acetone are the problems associated with hazardous waste disposal requirements, their flammability and noxiousness if vapors escape into the laboratory. Agents such as dimethyl sulfoxide, sugars, and amino acids which replace water in the cell, have also been used to preserve suspended cells, but these cells must be stored frozen and washed prior to use.
Whenever cells are stored frozen, there is the subsequent problem of breaking the cell membrane during freeze-thaw cycles and thus a consequent loss of cells. Additional problems associated with methods that involve freezing of the cells is the cost of maintaining freezers and the loss of laboratory space and the inconvenience of having to thaw and wash the cells prior to use.
Thus, there is a need in the field of immunology for a method for drying or fixing monolayers of mammalian cells to solid-phase supports for use in immunoassays which obviate the problems discussed above.
It is therefore a primary object of the present invention to provide a method for drying monolayers of mammalian cells adsorbed onto a solid-phase support for use in immunoassays in a manner maintaining the complete antigenicity of the cell surface antigens and in a manner permitting the solid-phase supports having these dried cell monolayers thereon to be stored at ambient temperatures thereby avoiding the problems of storage at sub-freezing temperatures.
It is another object of the present invention to provide articles for use in solid-phase immunoassays where the mammalian cells dried thereon can be stored at room temperature for at least six months and remain capable of performing sensitive and accurate immunoassays.
It is a further object of the present invention to provide a method for drying monolayers of mammalian cells adsorbed onto a solid-phase support for use in immunoassays that reduces the costs involved in such assays by extending the shelf-life of the cell monolayer.
It is still another object of the present invention to provide a method for drying monolayers of mammalian cells adsorbed onto a solid-phase support for use in immunoassays where the drying solution is not harmful to the monolayer nor interferes with performance of the assay.
It is an aim of the present invention to provide a method for drying monolayers of mammalian cells such as erythrocytes, lymphocytes, leukocytes and platelets onto a solid-phase support for use in immunoassays where the drying solution comprises an aqueous solution of a monosaccharide disaccharide, trisaccharide or cyclitol and a salt.
Other and further objects will become apparent from the following description and claims.